Microfluidic devices continue to be of great interest for conducting analyses of chemical and biological analytes. The terms “microfluidic” or “microscale” device generally refer to devices for manipulating fluids that comprise a network of microfluidic elements (e.g., channels and/or chambers), in which at least one element has at least one dimension in the range of from about 0.5 μm to about 500 μm. For example, channels may have a depth and/or a width in this range, while a chamber may have at least a depth in this range.
Microfluidic devices enable small-scale reactions, which provide numerous benefits, such as reduced reagent usage, reduced sample size, and rapid operation, as is well known in the art. In addition, the integration of several functions within a single device is possible, wherein a sample may be transported from one device element to another for subsequent handling, reaction, or analysis. This aspect of integration in turn further enables improvements in sample throughput because of reduced sample handling by operators or robotic stations, smaller space requirements, and even portability for remote or field usage.
Moreover, numerous sample wells, reaction chambers, and/or analysis regions can be provided on a single device, which in principle allows numerous analyses to be conducted in parallel. Bottlenecks may nonetheless arise if processes can only be conducted in sequence. For example, sample throughput is often limited by an analysis step, as one sample must wait for the analysis of a previous sample to conclude where the detector system (e.g., laser or optical detector) or analysis channel is shared. The microfluidic device layout of the microfluidic channels and wells is also a consideration, and the geometry of the device can be used to improve throughput by reducing transit times for analytes from one microfluidic element to another.
An example of improved device throughput based on channel geometry was disclosed by Dubrow et al. in U.S. Pat. No. 5,976,336. Dubrow et al. disclosed devices for electrophoretic separation analysis of different samples provided in multiple sample reservoirs. The devices have a single analysis channel, but sample reservoirs are located on each side of the analysis channel to maximize the number of reservoirs while minimizing the distance samples must travel to reach the analysis channel. Furthermore, while one sample introduced from a reservoir on one side of the separation channel is being analyzed by electrophoretic separation, a sample from a reservoir on the opposite side of the channel can be preloaded, that is, brought to a position in the load channel close to the separation channel. Once the analysis of the first sample is finished, the sample prepositioned in the load channel can complete its transit along the load channel from its sample reservoir to the intersection with the analysis channel and then be injected into the analysis channel for analysis.
Devices comprising functions and microfluidic elements other than just multiple sample reservoirs present different challenges and bottlenecks to high-throughput and/or reduced analysis time. In particular, microfluidic devices comprising reaction chambers present a challenge for coordinating sample movement within the device with the progress of the reaction. For example, for thermocycled reactions the temperature-induced changes to the sample and/or the transport properties caused by thermocycling must be considered and will limit when samples can be removed for analysis or other uses.
One application well-suited to a microfluidic device is an integrated device that performs a nucleic acid amplification reaction and analyzes the amplification products. For example, combining a chamber for an amplification reaction, such as PCR, with a separation channel for capillary electrophoresis (CE) detection provides a method for following the progress of the amplification in real time and quantifying the amount of target in the original sample.
Examples of such integrated devices are known in the art, including PCR-CE devices. One such device is disclosed in U.S. Pat. No. 8,394,324, by Bousse and Zhang, assigned to the same assignee, which is herein incorporated by reference in its entirety. A solution in a reaction chamber in the device is thermocycled to generate PCR amplification products, and after certain cycles, a sample of the reaction product is removed and analyzed for the amount of product generated in the amplification reaction. Generally, however, PCR-CE integrated devices pause the thermocycling process while the CE separation is conducted, or at least until the sample is transported from the chamber to the separation channel. This pause lengthens the total time needed to complete the analysis. Accordingly, there remains a need for devices and methods that are capable of more efficiently coordinating sample movement and completing an analysis in less time in order to increase sample throughput and decrease operating costs.